Among others, HGF is a factor for supporting wound healing and is a protein expressed in the mesenchymal cells such as lung macrophages and fibroblasts, kupffer cells in the liver, and leukocytes. HGF is a cytokine, which is secreted at cell damage and appears to have an importance for the regeneration of certain organs and for the healing of wounds. Chemically HGF is a glycoprotein, which first is synthesized as a native (inactive) precursor. The precursor is cleaved to active HGF in the damaged organ via a particular activator. HGF and other factors bind to heparin, which seems to be important for the activation of HGF and the binding to its receptor. The receptor binding to HGF is c-MET. Since the c-MET receptor only is down regulated in damaged organs, it is only cells in these damaged organs that appear to respond to a HGF-receptor interaction. Examples of such factors in the growth factor family having heparin binding affinities which factors have an influence on the wound healing process, include by others Platelet derived growth factor (PDGF), Epidermal growth factor (EGF), Transforming growth factor alfa (TGF-α), Transforming growth factor beta (TGF-β), insulin like growth factor (IGF-I) and Fibroblast growth factor (FGF).
Antithrombin III (AT-III) is a plasma glycoprotein that inhibits serine proteases in the coagulation cascade and, thus, plays a major role in the regulation of blood clotting. AT-III is an inhibitor of Factors IXa, Xa, XI, XIIa, and thrombin. Thus, AT-III regulates clot formation in different stages of the coagulation cascade. A small decrease of AT-III content in the blood is associated with increased risk of thromboembolism. AT-III concentrates are used in the prophylaxis and treatment of thromboembolic disorders in patients with acquired or hereditary AT-III deficiency. In addition, it has been reported that AT-III is involved in many other biological responses, for example angiogenesis and inflammatory responses. The function of AT-III in these mechanisms is not yet fully understood.
Purification of AT-III with affinity chromatography, using heparin as the solid phase bound ligand, is known in the art. Miller-Andersson et al. (Thrombosis Research 5, 439-452, 1974) discloses the use of heparin-Sepharose to purify human AT-III. The entire procedure, which included ion exchange and gel filtration chromatography, provided a 34% yield.
Histidine-rich glycoprotein (HRGP) is a single-chained plasma protein originally isolated in 1972. The exact physiological function of HRGP is still unknown. Due to interaction with heparin, fibrinogen and fibrin, plasminogen and activated platelets, HRGP is considered to be a modulator of coagulation and fibrinolysis (Koide, T. In: Fibrinolysis: Current Prospects. Gaffney, P J (Ed.), John Libbey & Co., London 1988, p. 55-63). The polypeptide chain consists of 507 amino acid residues and contains regions that share homology with other plasma proteins, e.g. AT-III (Koide, T. et al. (1986) Biochemistry 25, 2220-2225).